Genetic Diversity And Phylogeny Of Rhizobia Isolated From Medicago And Lathyrus In The West Plateau Of Sichuan, China

Twenty-five rhizobial strains isolated from root nodules of Medicago and Lathyrus in the west plateau of Sichuan, were investigated by the fingerprinting16S rDNA PCR-RFLP, IGS rDNA PCR-RFLP,16S rDNA sequences, recA and atpD gene.The results demonstrated that a high biological diversity existed among these rhizobia of Medicago and Lathyrus which collected from Aba and Ganzi in Sichuan.The results of16S rDNA PCR-RFLP indicated that all of the tested and reference strains were clustered into2groups at the similarity of68%, and clustered into5groups at the similarity of85%, Sinorhizobium, Agrobacterium, Rhizobium and Mesorhizobium reference strains could be completely separated. Among them, the group1composed by4tested strains (such as SCAU626) and Sinorhizobium reference strains. The group2composed by10tested strains (such as SCAU627, SCAU713) and Agrobacterium reference strains. The group3composed by6tested strains (such as SCAU632> SCAU718) and Rhizobium reference strains. The group4composed by5tested strains, such as SCAU633> SCAU736. The group5are all composed of reference strains belongs to Mesorhizobium.The relustes of IGS rDNA PCR-RFLP analysis indicated that all of the tested and reference strains were clustered into2groups at the similarity of66%, and clustered into7genetic groups at the similarity level of87%. The reference strain of Sinorhizobium, Agrobacterium, Rhizobium and Mesorhizobium also could be completely separated at the similarity of78%. Among them, the group1composed by4tested strains (such as SCAU626) and Sinorhizobium reference strains. The group2composed by2tested strains (such as SCAU627).The group3composed by3tested strains (such as SCAU631、SCAU717) and Agrobacterium reference strains. The group4composed by5tested strains(such as SCAU731、SCAU713) and Rhizobium reference strains. The group5composed by6tested strains (such as SCAU632, SCAU644> SCAU718) and Rhizobium reference strains. The group6composed by5tested strains, such as SCAU633、SCAU736. The group7were all composed of reference strains belonged to Mesorhizobium. The results of16S-IGS rDNA PCR-RFLP showed that, With16S rDNA PCR-RFLP, IGS rDNA PCR-RFLP in reflecting the genus level differences, in order to obtain more accurate information, also need to16S rDNA sequence determination and the establishment of the phylogenetic tree.The results of16S-IGS rDNA PCR-RFLP showed that with the16S rDNA PCR-RFLP, IGS rDNA PCR-RFLP reflected the differences in the genus level. In order to obtain more accurate information,16S rDNA sequence determination and establishment of the phylogenetic tree were needed.As the results of16S、IGS rDNA PCR-RFLP analysis, six representavive strian were selected to do the sequencing. The results of the sequencing indicated that the representative strain SCAU718and SCAU637were in Rhizobium genus, SCAU713and SCAU717belonged to Agrobacterium genus. The representative strain SCAU629was in Sinorhizobium genus, while SCAU736was in Ochrobactrum genus.The results of housekeeping gene recA and atpD sequence analysis showed that the phylogenetic tree of housekeeping gene and16S rRNA gene sequence of tested strains were consistent. But the location and position of reference strains in the evolutionary tree of housekeeping gene and16S rRNA gene was different, indicating that the phylogenetic relationships of species with the genus were difference, suggested that there were gene transfer and gene recombination in the same genera.